Techniques to improve polyurethane membranes for implantable glucose sensors

ABSTRACT

The invention provides an implantable membrane for regulating the transport of analytes therethrough that includes a matrix including a first polymer; and a second polymer dispersed throughout the matrix, wherein the second polymer forms a network of microdomains which when hydrated are not observable using photomicroscopy at 400× magnification or less. In one aspect, the homogeneous membrane of the present invention has hydrophilic domains dispersed substantially throughout a hydrophobic matrix to provide an optimum balance between oxygen and glucose transport to an electrochemical glucose sensor.

FIELD OF THE INVENTION

[0001] The present invention relates generally to membranes for use incombination with implantable devices for evaluating an analyte in a bodyfluid. More particularly, the invention relates to membranes forcontrolling the diffusion of glucose therethrough to a glucose sensor.

BACKGROUND OF THE INVENTION

[0002] A biosensor is a device that uses biological recognitionproperties for the selective analysis of various analytes orbiomolecules. Generally, the sensor will produce a signal that isquantitatively related to the concentration of the analyte. Inparticular, a great deal of research has been directed toward thedevelopment of a glucose sensor that would function in vivo to monitor apatient's blood glucose level. Such a glucose sensor is useful in thetreatment of diabetes mellitus. In particular, an implantable glucosesensor that would continuously monitor the patient's blood glucose levelwould provide a physician with more accurate information in order todevelop optimal therapy. One type of glucose sensor is the amperometricelectrochemical glucose sensor. Typically, an electrochemical glucosesensor employs the use of a glucose oxidase enzyme to catalyze thereaction between glucose and oxygen and subsequently generate anelectrical signal. The reaction catalyzed by glucose oxidase yieldsgluconic acid and hydrogen peroxide as shown in the reaction below(equation 1):${{glucose} + O_{2}}\overset{glucose}{\underset{oxidase}{\rightarrow}}{{{gluconic}\quad {acid}} + {H_{2}O_{2}}}$

[0003] The hydrogen peroxide reacts electrochemically as shown below inequation 2:

H₂O₂→2H⁺+O₂+2e⁻

[0004] The current measured by the sensor is generated by the oxidationof the hydrogen peroxide at a platinum working electrode. According toequation 1, if there is excess oxygen for equation 1, then the hydrogenperoxide is stoichiometrically related to the amount of glucose thatreacts with the enzyme. In this instance, the ultimate current is alsoproportional to the amount of glucose that reacts with the enzyme.However, if there is insufficient oxygen for all of the glucose to reactwith the enzyme, then the current will be proportional to the oxygenconcentration, not the glucose concentration. For the glucose sensor tobe useful, glucose must be the limiting reagent, i.e., the oxygenconcentration must be in excess for all potential glucoseconcentrations. Unfortunately, this requirement is not easily achieved.For example, in the subcutaneous tissue the concentration of oxygen ismuch less that of glucose. As a consequence, oxygen can become alimiting reactant, giving rise to a problem with oxygen deficit.Attempts have been made to circumvent this problem in order to allow thesensor to continuously operate in an environment with an excess ofoxygen.

[0005] Several attempts have been made to use membranes of various typesin an effort to control the diffusion of oxygen and glucose to thesensing elements of glucose oxidase-based glucose sensors. One approachhas been to develop homogenous membranes having hydrophilic domainsdispersed substantially throughout a hydrophobic matrix to circumventthe oxygen deficit problem, where glucose diffusion is facilitated bythe hydrophilic segments.

[0006] For example, U.S. Pat. No. 5,322,063 to Allen et al. teaches thatvarious compositions of hydrophilic polyurethanes can be used in orderto control the ratios of the diffusion coefficients of oxygen to glucosein an implantable glucose sensor. In particular, various polyurethanecompositions were synthesized that were capable of absorbing from 10 to50% of their dry weight of water. The polyurethanes were renderedhydrophilic by incorporating polyethyleneoxide as their soft segmentdiols. One disadvantage of this invention is that the primary backbonestructure of the polyurethane is sufficiently different so that morethan one casting solvent may be required to fabricate the membranes.This reduces the ease with which the membranes may be manufactured andmay further reduce the reproducibility of the membrane. Furthermore,neither the percent of the polyethyleneoxide soft segment nor thepercent water pickup of the polyurethanes disclosed by Allen directlycorrelate to the oxygen to glucose permeability ratios. Therefore, oneskilled in the art cannot simply change the polymer composition and beable to predict the oxygen to glucose permeability ratios. As a result,a large number of polymers would need to be synthesized before a desiredspecific oxygen to glucose permeability ratio could be obtained.

[0007] U.S. Pat. Nos. 5,777,060 and 5,882,494, each to Van Antwerp, alsodisclose homogeneous membranes having hydrophilic domains dispersedthroughout a hydrophobic matrix to reduce the amount of glucosediffusion to the working electrode of a biosensor. For example, U.S.Pat. No. 5,882,494 to Van Antwerp discloses a membrane including thereaction products of a diisocyanate, a hydrophilic diol or diamine, anda silicone material. In addition, U.S. Pat. No. 5,777,060 to Van Antwerpdiscloses polymeric membranes that can be prepared from (a) adiisocyanate, (b) a hydrophilic polymer, (c) a siloxane polymer havingfunctional groups at the chain termini, and optionally (d) a chainextender. Polymerization of these membranes typically requires heatingof the reaction mixture for periods of time from 1 to 4 hours, dependingon whether polymerization of the reactants is carried out in bulk or ina solvent system. Therefore, it would be beneficial to provide a methodof preparing a homogenous membrane from commercial polymers. Moreover,as mentioned above, one skilled in the art cannot simply change thepolymer composition and be able to predict the oxygen to glucosepermeability ratios. Therefore, a large number of polymers would need tobe synthesized and coating or casting techniques optimized before adesired specific oxygen to glucose permeability ratio could be obtained.

[0008] A further membrane is disclosed in U.S. Pat. No. 6,200,772 B1 toVadgama et al. that has hydrophilic domains dispersed substantiallythroughout a hydrophobic matrix for limiting the amount of glucosediffusing to a working electrode. In particular, the patent describes asensor device that includes a membrane comprised of modifiedpolyurethane that is substantially non-porous and incorporates anon-ionic surfactant as a modifier. The non-ionic surfactant isdisclosed as preferably including a poly-oxyalkylene chain, such as onederived from multiple units of poly-oxyethylene groups. As described,the non-ionic surfactant may be incorporated into the polyurethane byadmixture or through compounding to distribute it throughout thepolyurethane. The non-ionic surfactant is, according to thespecification, preferably incorporated into the polyurethane by allowingit to react chemically with the polyurethane so that it becomeschemically bound into its molecular structure. Like most reactivepolymer resins, complete reaction of the surfactant into thepolyurethane may never occur. Therefore, a disadvantage of this membraneis that it can leach the surfactant over time and cause irritation atthe implant site or change its permeability to glucose.

[0009] PCT Application WO 92/13271 discloses an implantable fluidmeasuring device for determining the presence and the amounts ofsubstances in a biological fluid that includes a membrane for limitingthe amount of a substance that passes therethrough. In particular, thisapplication discloses a membrane including a blend of two substantiallysimilar polyurethane urea copolymers, one having a glucose permeabilitythat is somewhat higher than preferred and the other having a glucosepermeability that is somewhat lower than preferred.

[0010] An important factor in obtaining a useful implantable sensor fordetection of glucose or other analytes is the need for optimization ofmaterials and methods in order to obtain predictable in vitro and invivo function. The ability of the sensor to function in a predictableand reliable manner in vitro is dependent on consistent fabricationtechniques. Repeatability of fabrication has been a problem associatedwith prior art membranes that attempt to regulate the transport ofanalytes to the sensing elements.

[0011] We refer now to FIG. 1, which shows a photomicrograph at 200×magnification of a prior art cast polymer blend following hydration. Adisadvantage of the prior art membranes is that, upon thermodynamicseparation from the hydrophobic portions, the hydrophilic componentsform undesirable structures that appear circular 1 and elliptical 2 whenviewed with a light microscope when the membrane 3 is hydrated, but notwhen it is dry. These hydrated structures can be detected byphotomicroscopy under magnifications in the range of between 200×-400×,for example. They have been shown by the present inventors to benon-uniform in their dimensions throughout the membrane, with some beingof the same size and same order of dimensions as the electrode size. Itis believed that these large domains present a problem in that theyresult in a locally high concentration of either hydrophobic orhydrophilic material in association with the electrode. This can resultin glucose diffusion being limited or variable across the dimensionadjacent the sensing electrode. Moreover, these large hydratedstructures can severely limit the number of glucose diffusion pathsavailable. It is noted that particles 4 in membrane 3 are dustparticles.

[0012] With reference now to a schematic representation of a knownmembrane 14 in FIG. 2A, one can consider by way of example a continuouspath 16 by which glucose may traverse along the hydrophilic segments 10that are dispersed in hydrophobic sections 12 of the membrane. For path16, glucose is able to traverse a fairly continuous path along assembledhydrophilic segments 10 from the side 18 of the membrane in contact withthe body fluid containing glucose to the sensing side 20 proximal tosensor 22, where an electrode 24 is placed at position 26 where glucosediffusion occurs adjacent surface 20. In particular, in that portion ofthe membrane 14 proximal to position 26, glucose diffusion occurs alonghydrophilic segments 10 that comprise a hydrated structure 28 having asize and overall dimensions x that are of the same order of magnitude aselectrode 24. Therefore, glucose diffusion would be substantiallyconstant across the dimension adjacent electrode 24, but the number ofglucose diffusion paths would be limited.

[0013] Referring now to FIG. 2B, one can consider an example whereglucose traversing prior art membrane 14 from side 18 in contact withthe body fluid to the sensing side 20 cannot adequately reach electrode30. In particular, electrode 30 is located at position 34, which isadjacent to a locally high concentration of a hydrophobic region 12 ofprior art membrane 14. In this instance, glucose diffusion cannotadequately occur, or is severely limited across the dimension adjacentthe electrode surface. Consequently, one would expect that the locallyhigh concentration of the hydrophobic regions adjacent to workingelectrode 30 would limit the ability of the sensing device to obtainaccurate glucose measurements. The random chance that the membrane couldbe placed in the 2A configuration as opposed to 2B leads to widevariability in sensor performance.

[0014] We also refer to FIG. 2C, which shows another cross-section ofprior art membrane 14. In this instance, glucose is able to traverse afairly continuous path 36 from side 18 to side 20 proximal to thesensing device. However, electrode 38 is located at position 40 suchthat glucose diffusion is variable across the dimension adjacent theelectrical surface. In particular, most of the electrode surface isassociated with a locally high concentration of hydrophobic region and asmall portion is associated with hydrophilic segments 10 along glucosediffusion path 36. Furthermore, glucose diffusing along path 36 a wouldnot be associated with the electrode. Again, the large non-uniformstructures of the prior art membranes can limit the number of glucosediffusion paths and the ability of the sensing device to obtain accurateglucose measurements.

[0015] It would be beneficial to form more homogeneous membranes forcontrolling glucose transport from commercially available polymers thathave a similar backbone structure. This would result in a morereproducible membrane. In particular, it is desired that one would beable to predict the resulting glucose permeability of the resultingmembrane by simply varying the polymer composition. In this way, theglucose diffusion characteristics of the membrane could be modified,without greatly changing the manufacturing parameters for the membrane.In particular, there is a need for homogeneous membranes havinghydrophilic segments dispersed throughout a hydrophobic matrix that areeasy to fabricate reproducibly from readily available reagents. Ofparticular importance would be the development of membranes where thehydrophilic portions were distributed evenly throughout the membrane,and where their size and dimensions were on an order considerably lessthan the size and dimensions of the electrode of the sensing device toallow the electrode to be in association with a useful amount of bothhydrophobic and hydrophilic portions. The ability of the membranes to besynthesized and manufactured in reasonable quantities and at reasonableprices would be a further advantage.

SUMMARY OF THE INVENTION

[0016] The present invention provides an implantable membrane forcontrolling the diffusion of an analyte therethrough to a biosensor withwhich it is associated. In particular, the membrane of the presentinvention satisfies a need in the art by providing a homogenous membranewith both hydrophilic and hydrophobic regions to control the diffusionof glucose and oxygen to a biosensor, the membrane being fabricatedeasily and reproducibly from commercially available materials.

[0017] The invention provides a biocompatible membrane that regulatesthe transport of analytes that includes: (a) a matrix including a firstpolymer; and (b) a second polymer dispersed throughout the matrix,wherein the second polymer forms a network of microdomains which whenhydrated are not observable using photomicroscopy at 400× magnificationor less.

[0018] Further provided by the invention is a polymeric membrane forregulation of glucose and oxygen in a subcutaneous glucose measuringdevice that includes: (a) a matrix including a first polymer; and (b) asecond polymer dispersed throughout the matrix, wherein the secondpolymer forms a network of microdomains which are notphotomicroscopically observable when hydrated at 400× magnification orless.

[0019] Yet another aspect of the present invention is directed to apolymeric membrane for regulating the transport of analytes, themembrane including at least one block copolymer AB, wherein B forms anetwork of microdomains which are not photomicroscopically observablewhen hydrated at 400× magnification or less.

[0020] Also provided is a membrane and sensor combination, the sensorbeing adapted for evaluating an analyte within a body fluid, themembrane having: (a) a matrix including a first polymer; and (b) asecond polymer dispersed throughout the matrix, wherein the secondpolymer forms a network of microdomains which are notphotomicroscopically observable when hydrated at 400× magnification orless.

[0021] The invention further provides an implantable device formeasuring an analyte in a hydrophilic body fluid, including: (a) apolymeric membrane having (i) a matrix including a first polymer; and(ii) a second polymer dispersed throughout the matrix, wherein thesecond polymer forms a network of microdomains which are notphotomicroscopically observable when hydrated at 400× magnification orless; and (b) a proximal layer of enzyme reactive with the analyte.

[0022] Moreover, a method for preparing an implantable membraneaccording to the invention is provided, the method including the stepsof: (a) forming a composition including a dispersion of a second polymerwithin a matrix of a first polymer, the dispersion forming a network ofmicrodomains which are not photomicroscopically observable when hydratedat 400× magnification or less; (b) maintaining the composition at atemperature sufficient to maintain the first polymer and the secondpolymer substantially soluble; (c) applying the composition at thistemperature to a substrate to form a film thereon; and (d) permittingthe resultant film to dry to form the membrane.

BRIEF DESCRIPTION OF THE DRAWINGS

[0023]FIG. 1 is a photomicrograph of a cross-section of prior artmembrane at 200× magnification following hydration with water for twohours.

[0024]FIG. 2A is a schematic representation of a cross-section of aprior art membrane having large hydrated structures dispersedsubstantially throughout a hydrophobic matrix, the hydrated structuresbeing photomicroscopically observable at 400× magnification or less. Thefigure illustrates the positioning of a working electrode relative to aglucose diffusion pathway.

[0025]FIG. 2B is another schematic representation of a cross-section ofthe prior art membrane of FIG. 2A, where the working electrode is placedin association with a locally high concentration of the hydrophobicmatrix.

[0026]FIG. 2C is yet another schematic representation of a cross-sectionof the prior art membrane of FIG. 2A where glucose diffusion is variableacross the dimension adjacent the electrode surface.

[0027]FIG. 3 is a photomicrograph of a cross-section of a membrane ofthe present invention at 200× magnification following hydration withwater for two hours.

[0028]FIG. 4 is a schematic representation of a cross-sectionillustrating one particular form of the membrane of the presentinvention that shows a network of microdomains which are notphotomicroscopically observable at 400× or less magnification dispersedthrough a hydrophobic matrix, where the membrane is positioned inassociation with a sensor that includes a working electrode.

[0029]FIG. 5 is a schematic representation of a cross-section of themembrane of FIG. 3 in combination with an enzyme containing layerpositioned more adjacent to a sensor 50.

[0030]FIG. 6 is a graph showing sensor output versus the percent of thehydrophobic-hydrophilic copolymer component in the coating blend.

[0031]FIG. 7 is a graph showing the percent standard deviation of thesensor current versus the percent of the hydrophobic-hydrophiliccopolymer component in the coating blend.

DETAILED WRITTEN DESCRIPTION

[0032] In order to facilitate understanding of the present invention, anumber of terms are defined below.

[0033] The term “analyte” refers to a substance or chemical constituentin a biological fluid (e.g. blood or urine) that is intended to beanalyzed. A preferred analyte for measurement by analyte detectingdevices including the membrane of the present invention is glucose.

[0034] The term “sensor” refers to the component or region of a deviceby which an analyte can be evaluated.

[0035] By the terms “evaluated”, “monitored”, “analyzed”, and the like,it is meant that an analyte may be detected and/or measured.

[0036] The phrase “continuous glucose sensing” refers to the period inwhich monitoring of plasma glucose concentration is repeatedly performedover short periods of time, for example, 10 seconds to about every 15minutes.

[0037] The term “domain” refers to regions of the membrane of thepresent invention that may be layers, uniform or non-uniform gradients(e.g. anisotropic) or provided as portions of the membrane. Furthermore,the region possesses physical properties distinctly different from otherportions of the membrane.

[0038] The terms “accurate” and “accurately” means, for example, 85% ofmeasured glucose values are within the “A” and “B” region of a standardClarke Error Grid when the sensor measurements are compared to astandard reference measurement. It is understood that like anyanalytical device, calibration, calibration validation and recalibrationare required for the most accurate operation of the device.

[0039] The term “host” refers to humans and other animals.

[0040] In the disclosure that follows, the invention will primarily bereferred to in terms of assay of glucose and solutions such as bloodthat tend to contain a large excess of glucose over oxygen. However, itis well within the contemplation of the present invention that themembrane is not limited solely to the assay of glucose in a biologicalfluid, but may be used for the assay of other compounds. In addition,the sensor primarily referred to is an electrochemical sensor thatdirectly measures hydrogen peroxide. However, it is well within thecontemplation of the present invention that non-electrochemical basedsensors that use optical detectors or other suitable detectors may beused to evaluate an analyte.

[0041] Membranes of the prior art have generally been unreliable atlimiting the passage of glucose to implantable glucose sensors. This haspresented a problem in the past in that the amount of glucose cominginto contact with the immobilized enzyme exceeds the amount of oxygenavailable. As a result, the oxygen concentration is the rate-limitingcomponent of the reaction, rather than the glucose concentration, suchthat the accuracy of the glucose measurement in the body fluid iscompromised.

[0042] As described above, in contrast to the present invention, adisadvantage of prior art membranes for regulating analyte transporttherethrough has been their tendency to form large undesirablestructures (see FIG. 1) that are observable when the membrane ishydrated. In particular, these hydrated structures can be detected byphotomicroscopy under magnifications in the range of between 200×-400×,for example. They have been shown by the present inventors to benon-uniform in their dimensions through the membrane, with some being ofthe same size and same order of dimensions as the electrode size. Theselarge structures have been found to be problematic in that they canresult in a locally high concentration of either hydrophobic orhydrophilic material in association with the working electrode, whichcan lead to inaccurate glucose readings. Moreover, they can greatlyreduce the number of glucose diffusion paths available.

[0043] The membrane of the present invention seeks to circumvent theseproblems associated with prior art membranes by providing a reliablehomogeneous membrane that regulates the transport of glucose or otheranalytes therethrough, the membrane having (a) a matrix including afirst polymer; and (b) a second polymer dispersed throughout the matrix,wherein the second polymer forms a network of microdomains which whenhydrated are not observable using photomicroscopy at 400× magnificationor less. In one embodiment of the invention, the membrane issubstantially free of observable domains.

[0044] We refer now to FIG. 3, which shows a photomicrograph of across-section of a membrane 5 according to the present inventionfollowing hydration at two hours. As shown in FIG. 3, the membrane isdevoid of any undesirable, large elliptical or spherical structures,such as were observable in hydrated prior art membranes at similarmagnifications. It is noted that particles 6 in membrane 5 are dustparticles.

[0045] For purposes of the present invention, it is likely that glucosepermeability and diffusion is related to the ratio of hydrophobic tohydrophilic constituents and their distribution throughout the membrane,with diffusion occurring substantially along assembled hydrophilicsegments from the side of the membrane in contact with the host to thesensing side.

[0046] Referring now to FIG. 4, membrane 42 of the present invention, inaccordance with a particular arrangement, is schematically shown havinghydrophilic segments 44 dispersed substantially throughout a hydrophobicmatrix 46 and presenting a surface 48 to a hydrophilic body fluid. Thehydrophilic body fluid contains the sample to be assayed. In oneembodiment, the body fluid contains both glucose and oxygen. Membrane 42restricts the rate at which glucose enters and passes through themembrane and/or may increase the rate at which oxygen enters and passesthrough membrane 42.

[0047] While not wishing to be bound by any one theory, it is likelythat glucose diffuses substantially along hydrophilic segments 44, butis generally excluded from the hydrophobic matrix 46. It is noted thatwhile the hydrophilic segments 44 are shown as comprising discretemicrodomains in FIG. 4, small amounts of hydrophobic polymer may bepresent therein, particularly at the interface with the hydrophobicmatrix 46. Similarly, small amounts of hydrophilic polymer may bepresent in the hydrophobic matrix 46, particularly at the interface withhydrophilic segments 44.

[0048] In the embodiment shown in FIG. 4, inventive membrane 42 is shownin combination with a sensor 50, which is positioned adjacent to themembrane. It is noted that additional membranes or layers may besituated between membrane 42 and sensor 50, as will be discussed infurther detail below. Diffusion of the sample along paths 52 throughmembrane 42 into association with a working electrode 54 of sensor 50causes development of a signal that is proportional to the amount ofanalyte in the sample. Determination of the analyte may be made bycalculations based upon similar measurements made on standard solutionscontaining known concentrations of the analyte. For example, one or moreelectrodes may be used to detect the amount of analyte in the sample andconvert that information into a signal; the signal may then betransmitted to electronic circuitry required to process biologicalinformation obtained from the host. U.S. Pat. Nos. 4,757,022, 5,497,772and 4,787,398 describe suitable electronic circuitry that may beutilized with implantable devices of the present invention.

[0049] The present invention solves a need in the art by providing areliable membrane for controlling glucose diffusion therethrough. Asshown in FIG. 4, glucose can traverse along hydrophilic segments 44 fromthe side 48 of the membrane in contact with a body fluid to the side 56proximal to sensor 50. The hydrophilic microdomains 44 are likelydistributed substantially evenly throughout the membrane. Furthermore,these microdomains are likely substantially uniform in size throughoutthe membrane. The size and order to dimensions of these microdomains isconsiderably less than the that of the working electrode 54 of sensor50. As such, the electrode is in association with a useful amount ofboth the hydrophobic 46 and hydrophilic 44 regions of the membrane toallow effective control over the amount of glucose diffusing to theelectrode. Moreover, as shown in FIG. 4, the number of paths availablefor glucose to permeate the membrane and diffuse from side 48 to thesensing side 56 would be greater for the inventive membrane than forprior art membranes. Consequently, more accurate and reproducibleglucose readings are attainable across the entire inventive membrane.

[0050]FIG. 5 shows a preferred embodiment of the present inventionwherein membrane 42 is used in combination with a proximal membranelayer 58 that comprises an enzyme that is reactive with the analyte. Inthis instance, diffusion of the sample from side 48 through the membrane42 into contact with the immobilized enzyme in layer 58 leads to anenzymatic reaction in which the reaction products may be measured. Forexample, in one embodiment the analyte is glucose. In a furtherembodiment, the enzyme immobilized in layer 58 is glucose oxidase.

[0051] As described above, glucose oxidase catalyzes the conversion ofoxygen and glucose to hydrogen peroxide and gluconic acid. Because foreach glucose molecule metabolized, there is proportional change in theco-reactant O₂ and the product H₂O₂, one can monitor the change ineither the co-reactant or the product to determine glucoseconcentration. With further reference to FIG. 5, diffusion of theresulting hydrogen peroxide through layer 58 to the sensor 50, (e.g.electrochemically reactive surfaces), causes the development of anelectrical current that can be detected. This enables determination ofthe glucose by calculations based upon similar measurements made onstandard solutions containing known concentrations of glucose.

[0052] In addition to glucose oxidase, the present inventioncontemplates the use of a layer impregnated with other oxidases, e.g.galactose oxidase or uricase. For an enzyme-based electrochemicalglucose sensor to perform well, the sensor's response must neither belimited by enzyme activity nor cofactor concentration. Because enzymes,including glucose oxidase, are subject to deactivation as a function ofambient conditions, this behavior needs to be accounted for inconstructing sensors for long-term use.

[0053] When the membrane of the present invention is combined with anenzyme layer 58 as shown in FIG. 5, it is the enzyme layer that islocated more proximally to the sensor 50 (e.g. electrochemicallyreactive surfaces). It is noted that enzyme-containing layer 58 must beof sufficient permeability to 1) freely pass glucose to active enzymeand 2) to permit the rapid passage of hydrogen peroxide to the sensor(electrode surface). A failure to permit the rapid passage of glucose tothe active enzyme or hydrogen peroxide from the active enzyme to theelectrode surface can cause a time delay in the measured signal andthereby lead to inaccurate results.

[0054] Preferably, the enzyme layer is comprised of aqueouspolyurethane-based latex into which the enzyme is immobilized.

[0055] It is noted that while the inventive membrane 42 may itselfcontain immobilized enzymes for promoting a reaction between glucose andoxygen, it is preferred that the enzyme be located in a separate layer,such as layer 58 shown in FIG. 5. As described above, it is known thatenzyme actively reacting with glucose is more susceptible toirreversible inactivation. Therefore, a disadvantage of providing enzymein a layer that is semi-permeable to glucose, is that the calibrationfactors of the sensor may change over time as the working enzymedegrades. In contrast, when enzyme is dispersed throughout a membranefreely permeable to glucose (i.e. layer 58 in FIG. 5), such a membraneis likely to yield calibration factors that are more stable over thelife of a sensor.

[0056] In one preferred embodiment of the invention, the first polymerof the membrane includes homopolymer A and the second polymer includescopolymer AB.

[0057] In another embodiment, the first polymer includes copolymer ABand the second polymer includes copolymer AB. Preferably, the amount ofB in copolymer AB of the first polymer is different than the amount of Bin copolymer AB of the second polymer. In particular, the membrane maybe formed from a blend of two AB copolymers, where one of the copolymerscontains more of a hydrophilic B polymer component than the blendedtargeted amount and the other copolymer contains less of a hydrophilic Bpolymer component than the blended targeted amount.

[0058] In yet another embodiment of the invention, the first polymerincludes homopolymer A and the second polymer includes homopolymer B.

[0059] As described above, the invention also provides a polymericmembrane for regulating the transport of analytes that includes at leastone block copolymer AB, wherein B forms a network of microdomains whichare not photomicroscopically observable when hydrated at 400×magnification or less. In one embodiment, the ratio of A to B incopolymer AB is 70:30 to 90:10.

[0060] For each of the inventive embodiments herein described,homopolymer A is preferably a hydrophobic A polymer. Moreover, copolymerAB is preferably a hydrophobic-hydrophilic copolymer component thatincludes the reaction products of a hydrophobic A polymer and ahydrophilic B polymer. Suitable materials for preparing membranes thepresent invention are described below.

[0061] For purposes of the present invention, copolymer AB may be arandom or ordered block copolymer. Specifically, the random or orderedblock copolymer may be selected from the following: ABA block copolymer,BAB block copolymer, AB random alternating block copolymer, AB regularlyalternating block copolymer and combinations thereof.

[0062] In a preferred embodiment, the sensor, membrane, and methods ofthe present invention may be used to determine the level of glucose orother analytes in a host. The level of glucose is a particularlyimportant measurement for individuals having diabetes in that effectivetreatment depends on the accuracy of this measurement.

[0063] In particular, the invention provides a method of measuringglucose in a biological fluid that includes the steps of: (a) providing(i) a host, and (ii) an implantable device for measuring an analyte in ahydrophilic body fluid, where the device includes a polymeric membranehaving a matrix including a first polymer and a second polymer dispersedthroughout the matrix, wherein the second polymer forms a network ofmicrodomains which are not photomicroscopically observable when hydratedat 400× magnification or less; and a proximal layer of enzyme reactivewith the analyte; and (b) implanting the device in the host. In oneembodiment, the device is implanted subcutaneously.

[0064] The invention also provides a method of measuring glucose in abiological fluid that includes the following steps: (a) providing (i) ahost, and (ii) an implantable device for measuring an analyte in ahydrophilic body fluid, that includes a polymeric membrane including amatrix including a first polymer and a second polymer dispersedthroughout the matrix, wherein the second polymer forms a network ofmicrodomains which are not photomicroscopically observable when hydratedat 400× magnification or less; and a proximal layer of enzyme reactivewith the analyte, the device being capable of accurate continuousglucose sensing; and (b) implanting the device in the host. Desirably,the implant is placed subcutaneously in the host.

[0065] Glucose sensors that use, for example, glucose oxidase to effecta reaction of glucose and oxygen are known in the art, and are withinthe skill of one in the art to fabricate (see, for example, U.S. Pat.Nos. 5,165,407, 4,890,620, 5,390,671, 5,391,250, 6,001,067 as well ascopending, commonly owned U.S. patent application Ser. No. 09/916,858.It is noted that the present invention does not depend on a particularconfiguration of the sensor, but is rather dependent on the use of theinventive membrane to cover or encapsulate the sensor elements.

[0066] For the electrochemical glucose sensor to provide useful results,the glucose concentration, as opposed to oxygen concentration, must bethe limiting factor. In order to make the system sensitive to glucoseconcentration, oxygen must be present within the membrane in excess ofthe glucose. In addition, the oxygen must be in sufficient excess sothat it is also available for electrochemical reactions occurring at theamperometric electrode surfaces. In a preferred embodiment, theinventive membrane is designed so that oxygen can pass readily into andthrough the membrane and so that a reduced amount of glucose diffusesinto and through the membrane into contact with an immobilized glucoseoxidase enzyme. The inventive membrane allows the ratio of oxygen toglucose to be changed from a concentration ratio in the body fluid ofabout approximately 50 and 100 parts of glucose to 1 of oxygen to a newratio in which there is a stoichiometric excess of oxygen in the enzymelayer. Through the use of the inventive membrane, an implantable glucosesensor system is not limited by the concentration of oxygen present insubcutaneous tissues and can therefore operate under the premise thatthe glucose oxidase reaction behaves as a 1-substrate (glucose)dependent process.

[0067] The present invention provides a semi-permeable membrane thatcontrols the flux of oxygen and glucose to an underlying enzyme layer,rendering the necessary supply of oxygen in non-rate-limiting excess. Asa result, the upper limit of linearity of glucose measurement isextended to a much higher value than that which could be achievedwithout the membrane of the present invention. In particular, in oneembodiment the membrane of the present invention is a polymer membranewith oxygen-to-glucose permeability ratios of approximately 200:1; as aresult, 1-dimensional reactant diffusion is adequate to provide excessoxygen at all reasonable glucose and oxygen concentrations found in asubcutaneous matrix [Rhodes, et al., Anal. Chem., 66: 1520-1529 (1994)].

[0068] A hydrophilic or “water loving” solute such as glucose is readilypartitioned into a hydrophilic material, but is generally excluded froma hydrophobic material. However, oxygen can be soluble in bothhydrophilic and hydrophobic materials. These factors affect entry andtransport of components in the inventive membrane. The hydrophobicportions of the inventive membrane hinder the rate of entry of glucoseinto the membrane, and therefore to the proximal enzyme layer whileproviding access of oxygen through both the hydrophilic and hydrophobicportions to the underlying enzyme.

[0069] In one preferred embodiment, the membrane of the invention isformed from a blend of polymers including (i) a hydrophobic A polymercomponent; and (ii) a hydrophobic-hydrophilic copolymer componentblended with component (i) that forms hydrophilic B domains that controlthe diffusion of an analyte therethrough, wherein the copolymercomponent includes a random or ordered block copolymer. Suitable blockcopolymers are described above. One is able to modify the glucosepermeability and the glucose diffusion characteristics of the membraneby simply varying the polymer composition.

[0070] In one preferred embodiment, the hydrophobic A polymer is apolyurethane. In a most preferred embodiment, the polyurethane ispolyetherurethaneurea. A polyurethane is a polymer produced by thecondensation reaction of a diisocyanate and a difunctionalhydroxyl-containing material. A polyurethaneurea is a polymer producedby the condensation reaction of a diisocyanate and a difunctionalamine-containing material. Preferred diisocyanates include aliphaticdiisocyanates containing from 4 to 8 methylene units. Diisocyanatescontaining cycloaliphatic moieties, may also be useful in thepreparation of the polymer and copolymer components of the membrane ofthe present invention. The invention is not limited to the use ofpolyurethanes as the hydrophobic polymer A component. The material thatforms the basis of the hydrophobic matrix of the inventive membrane maybe any of those known in the art as appropriate for use as membranes insensor devices and having sufficient permeability to allow relevantcompounds to pass through it, for example, to allow an oxygen moleculeto pass through the inventive membrane from the sample under examinationin order to reach the active enzyme or electrochemical electrodes.Examples of materials which may be used to make a non-polyurethane typemembrane include vinyl polymers, polyethers, polyesters, polyamides,inorganic polymers such as polysiloxanes and polycarbosiloxanes, naturalpolymers such as cellulosic and protein based materials and mixtures orcombinations thereof.

[0071] As described above, the hydrophobic-hydrophilic copolymercomponent includes the reaction products of a hydrophobic A polymercomponent and a hydrophilic B polymer component. The hydrophilic Bpolymer component is desirably polyethylene oxide. For example, oneuseful hydrophobic-hydrophilic copolymer component is a polyurethanepolymer that includes about 20% hydrophilic polyethyelene oxide. Thepolyethylene oxide portion of the copolymer is thermodynamically drivento separate from the hydrophobic portions of the copolymer and thehydrophobic A polymer component. The 20% polyethylene oxide based softsegment portion of the copolymer used to form the final blend controlsthe water pick-up and subsequent glucose permeability of the membrane ofthe present invention.

[0072] The polyethylene oxide may have an average molecular weight offrom 200 to 3000 with a preferred molecular weight range of 600 to 1500and preferably constitutes about 20% by weight of the copolymercomponent used to form the membrane of the present invention.

[0073] It is desired that the membrane of the present invention have athickness of about 5 to about 100 microns. In preferred embodiments, themembrane of the present invention is constructed of apolyetherurethaneurea/polyetherurethaneurea-block-polyethylene glycolblend and has a thickness of not more than about 100 microns, morepreferably not less than about 10 microns, and not more than about 80microns, and most preferably, not less than about 20 microns, and notmore than about 60 microns.

[0074] The membrane of the present invention can be made by casting fromsolutions, optionally with inclusion of additives to modify theproperties and the resulting cast film or to facilitate the castingprocess.

[0075] The present invention provides a method for preparing theimplantable membrane of the invention. The method includes the steps of:(a) forming a composition including a dispersion of a second polymerwithin a matrix of a first polymer, the dispersion forming a network ofmicrodomains which are not photomicroscopically observable when hydratedat 400× magnification or less; (b) maintaining the composition at atemperature sufficient to maintain the first polymer and the secondpolymer substantially soluble; (c) applying the composition at thetemperature to a substrate to form a film thereon; and (d) permittingthe resultant film to dry to form the membrane. In one embodiment, theforming step includes forming a mixture or a blend. As described above,in preferred embodiments, the first polymer is a polyurethane and thesecond polymer is polyethylene oxide. In general, the second polymer maybe a random or ordered block copolymer selected from the following: ABAblock copolymer, BAB block copolymer, AB random alternating blockcopolymer, AB regularly alternating block copolymer and combinationsthereof.

[0076] In one embodiment, the composition comprised of a dispersion ofthe second polymer within the matrix of a first polymer is heated to atemperature of about 70° C. to maintain the first and second polymerssubstantially soluble. For example, the combination of a hydrophobicpolymer A component and a hydrophobic-hydrophilic copolymer AB componentis desirably exposed to a temperature of about 70° C. to maintain thepolymer and copolymers substantially soluble. In particular, the blendis heated well above room temperature in order to keep the hydrophilicand hydrophobic components soluble with each other and the solvent.

[0077] The invention contemplates permitting the coated film formed onthe substrate to dry at a temperature from about 120° C. to about 150°C. The elevated temperature further serves to drive the solvent from thecoating as quickly as possible. This inhibits the hydrophilic andhydrophobic portions of the membrane from segregating and forming largeundesired structures.

[0078] The membrane and sensor combinations of the present inventionprovide a significant advantage over the prior art in that they provideaccurate sensor operation at temperatures from about 30° C. to about 45°C. for a period of time exceeding about 30 days to exceeding about ayear.

EXAMPLES Example 1

[0079] A Method for Preparing a Membrane of the Present Invention

[0080] The inventive membrane may be cast from a coating solution. Thecoating solution is prepared by placing approximately 281 gm ofdimethylacetamide (DMAC) into a 3 L stainless steel bowl to which asolution of polyetherurethaneurea (344 gm of Chronothane H (CardiotechInternational, Inc., Woburn, Mass.), 29,750 cp @ 25% solids in DMAC) isadded. To this mixture is added another polyetherurethaneurea(approximately 312 gm, Chronothane 1020 (Cardiotech International, Inc.,Woburn, Mass.), 6275 cp @ 25% solids in DMAC). The bowl is then fittedto a planetary mixer with a paddle-type blade and the contents arestirred for 30 minutes at room temperature. Coatings solutions preparedin this manner are then coated at between room temperature to about 70°C. onto a PET release liner (Douglas Hansen Co., Inc., Minneapolis,Minn.) using a knife-over-roll set at a 0.012 inch gap. The film iscontinuously dried at 120° C. to about 150° C. The final film thicknessis approximately 0.0015 inches.

[0081] An Optical Method for Evaluating a Membrane of the PresentInvention

[0082] A ¼″ by ¼″ piece of membrane is first immersed in deionized waterfor a minimum of 2 hours at room temperature. After this time, thesample is placed onto a microscope slide along with one drop of water. Aglass cover slide is then placed over the membrane and gentle pressureis applied in order to remove excess liquid from underneath the coverglass. In this way, the membrane does not dry during its evaluation. Thehydrated membrane sample is first observed at 40×-magnification using alight microscope (Nikon Eclipse E400). If air bubbles are present on thetop or bottom of the film, the cover glass is gently pressed again witha tissue in order to remove them. Magnification is then increased to200×; and the hydrated membrane is continuously observed while changingthe focus from the top to bottom of the film. This is followed by anincrease in magnification to 400×, with the membrane again beingcontinuously observed while changing the focus from the top to bottom ofthe film.

[0083] Results

[0084] Based on the results of an optical micrograph of a samplemembrane prepared by using a room temperature coating solution anddrying of the coated film at 120° C., the micrograph being captured asdescribed above, it was noticed that both circular and ellipticaldomains were present throughout the hydrated section of membrane. At thesame magnification, the domains were not observable in dry membrane.Giving that in an electrochemical sensor, the electrodes includedtherein are typically of the same size and same order of dimensions asthe observed circular and elliptical domains, such domains are notdesired. These domains present a problem in that they result in alocally high concentration of either hydrophilic or hydrophobic materialin association with the electrodes.

Example 2

[0085] Optimizing the Coating Solution Conditions

[0086] This example demonstrates that preheating the coating solution toa temperature of 70° C. prior to coating eliminates the presence of boththe circular and elliptical domains that were present throughout thehydrated cross-section of a membrane prepared using a room temperaturecoating solution and drying of the coated film at 120° C. Example 2further demonstrates that, provided the coating solution is preheated toabout 70° C., either a standard (120°) or elevated (150° C.) dryingtemperature were sufficient to drive the DMAC solvent from the coatedfilm quickly to further inhibit the hydrophilic and hydrophobic portionsof the polyurethane membrane from segregating into large domains.

[0087] In particular, the invention was evaluated by performing acoating experiment where standard coating conditions (room temperaturecoating solution and 120° C. drying temperature of the coated film) werecompared to conditions where the coating solution temperature waselevated and/or the drying temperature of the coated film was elevated.Four experimental conditions were run as follows:

[0088] SS-room temperature solution and standard (120° C.) oventemperature.

[0089] SE-room temperature solution and elevated (150° C.) oventemperature.

[0090] ES-preheated (70° C.) solution and standard (120° C.) oventemperature.

[0091] EE-preheated (70° C.) solution and elevated (150° C.) oventemperature.

[0092] Results

[0093] Samples of each of the four membranes listed above were thenhydrated for 2 hours, and then observed under the microscope.Performance specifications were achieved when the micrograph of themembrane prepared under a given condition showed an absence of circularand/or elliptical domains that result in an undesirable, discontinuoushydrophilic and hydrophobic membrane structure. Table 1 below summarizesthese results where (+) indicates a membrane meeting desired performancespecifications and (−) is indicative of a membrane showing theundesirable circular and/or elliptical domains. In summary, for both theES and EE conditions, where the coating solution was preheated to 70° C.prior to coating on a substrate, no hydrated domains were observed at a200× magnification. Furthermore, regardless of the drying temperatureused for the coated film, when the coating solution was not preheated(conditions SS and SE), the hydrated structures were observed.Therefore, it is likely that preheating the coating solution effectivelyinhibits the hydrophilic and hydrophobic segments of the polyurethanefrom segregating into large domains. TABLE 1 Coating Condition Result SS− SE − ES (Inventive) + EE (Inventive) +

Example 3

[0094] Evaluation of the Inventive Membranes for Their Permeability toGlucose and H₂O₂

[0095] Membranes prepared under the EE condition described in Example 2were evaluated for their ability to allow glucose and hydrogen peroxideto get through the membrane to a sensor. In particular, a series ofpolyurethane blends of the present invention were generated wherein thepercentage of Chronothane H in a coating blend was varied. Furthermore,one of these blends (57.5% Chronothane H in coating blend) was preparedunder both the EE condition and the SS condition as described in Example2. FIG. 6 shows that the sensor output generated with a series ofpolyurethane blends of the present invention was dependent upon thepercentage of the Chronothane H. In particular, the sensor outputincreased as the percentage of Chronothane H in the coating blendincreased. With further reference to FIG. 6, when the percentage ofChronothane H in the coating blend was 57.5%, the sensor output wasthree times greater for the membrane prepared under the optimized EEcoating condition as compared to the non-optimized SS coating condition.

[0096] Furthermore, FIG. 7 demonstrates that, regardless of the percentChronothane H in the coating blend, an inventive membrane prepared underthe EE condition shows a fairly constant percent standard deviation ofsensor output. Moreover, a membrane prepared with 57.5% Chronothane H inthe coating blend under the SS condition showed a percent standarddeviation of sensor output approximately twice that of an EE membraneprepared with the same percentage of Chronothane H in the blend. It isnoted that given that the sensor output is a true measure of the amountof glucose getting through the membrane to the sensor, the resultsindicate that the permeability of glucose and H₂O₂ is relativelyconstant throughout a given inventive membrane prepared under optimizedcoating conditions (i.e., EE conditions). This is important from amanufacturing standpoint.

[0097] Having described the particular, preferred embodiments of theinvention herein, it should be appreciated that modifications may bemade therethrough without departing from the contemplated scope of theinvention. The true scope of the invention is set forth in the claimsappended hereto.

What is claimed is:
 1. A membrane that regulates the transport ofanalytes comprising: (a) a matrix comprising a first polymer; and (b) asecond polymer dispersed throughout said matrix, wherein said secondpolymer forms a network of microdomains which when hydrated are notobservable using photomicroscopy at 400× magnification or less.
 2. Themembrane of claim 1, wherein said membrane is substantially free ofobservable domains.
 3. The membrane of claim 1, wherein said firstpolymer comprises homopolymer A and said second polymer comprisescopolymer AB.
 4. The membrane of claim 3, wherein copolymer AB comprisesa random or ordered block copolymer.
 5. The membrane of claim 4, whereinsaid random or ordered block copolymer is selected from the groupconsisting of ABA block copolymer, BAB block copolymer, AB randomalternating block copolymer, AB regularly alternating block copolymerand combinations thereof.
 6. The membrane of claim 1, wherein said firstpolymer comprises copolymer AB and wherein said second polymer comprisescopolymer AB.
 7. The membrane of claim 6, wherein copolymer AB comprisesa random or ordered block copolymer.
 8. The membrane of claim 7, whereinsaid random or ordered block copolymer is selected from the groupconsisting of ABA block copolymer, BAB block copolymer, AB randomalternating block copolymer, AB regularly alternating block copolymerand combinations thereof.
 9. The membrane of claim 6, wherein the amountof B in copolymer AB of said first polymer is different than the amountof B in copolymer AB of said second polymer.
 10. The membrane of claim1, wherein said first polymer comprises homopolymer A and said secondpolymer comprises homopolymer B.
 11. The membrane of claim 1, whereinthe analyte is glucose.
 12. The membrane of claim 1, further includingan enzyme reactive with said analyte.
 13. The membrane of claim 12,wherein said enzyme is glucose oxidase.
 14. The membrane of claim 1, incombination with a proximal layer of enzyme reactive with said analyte.15. The membrane of claim 14, wherein said enzyme is glucose oxidase.16. The membrane of claim 1, wherein said second polymer is ahydrophobic-hydrophilic copolymer component comprising the reactionproducts of a hydrophobic A polymer and a hydrophilic B polymer.
 17. Themembrane of claim 16, wherein the hydrophilic B polymer is polyethyleneoxide.
 18. The membrane of claim 1, wherein the hydrophobic A polymer isa polyurethane.
 19. The membrane of claim 18, wherein the polyurethaneis polyetherurethaneurea.
 20. A polymeric membrane for regulation ofglucose and oxygen in a subcutaneous glucose measuring devicecomprising: (a) a matrix comprising a first polymer; and (b) a secondpolymer dispersed throughout said matrix, wherein said second polymerforms a network of microdomains which are not photomicroscopicallyobservable when hydrated at 400× magnification or less.
 21. A polymericmembrane for regulating the transport of analytes comprising at leastone block copolymer AB, wherein B forms a network of microdomains whichare not photomicroscopically observable when hydrated at 400×magnification or less.
 22. The membrane of claim 21, wherein the ratioof A to B in said copolymer is 70:30 to 90:10.
 23. A membrane and sensorcombination, said sensor being adapted for evaluating an analyte withina body fluid, said membrane comprising: (a) a matrix comprising a firstpolymer; and (b) a second polymer dispersed throughout said matrix,wherein said second polymer forms a network of microdomains which arenot photomicroscopically observable when hydrated at 400× magnificationor less.
 24. The combination of claim 23, wherein the sensor comprisesan electrochemical sensor.
 25. The combination of claim 23, wherein theanalyte diffuses through the membrane to the sensor for evaluationthereof.
 26. An implantable device for measuring an analyte in ahydrophilic body fluid, comprising: (a) a polymeric membrane comprising(i) a matrix comprising a first polymer; and (ii) a second polymerdispersed throughout said matrix, wherein said second polymer forms anetwork of microdomains which are not photomicroscopically observablewhen hydrated at 400× magnification or less; and (b) a proximal layer ofenzyme reactive with said analyte.
 27. The device of claim 26, furtherincluding a sensor for evaluating the analyte.
 28. The device of claim26, wherein the sensor comprises an electrochemical sensor.
 29. Thedevice of claim 26, wherein the analyte is glucose.
 30. The device ofclaim 26, wherein the enzyme is glucose oxidase.
 31. A method ofmonitoring glucose levels, comprising: (a) providing (i) a host, and(ii) an implantable device according to claim 26; and (b) implantingsaid device in said host.
 32. The method according to claim 31, whereinsaid implanting is subcutaneous.
 33. A method of measuring glucose in abiological fluid, comprising: (a) providing (i) a host, and (ii) animplantable device according to claim 26, said device being capable ofaccurate continuous glucose sensing; and (b) implanting said device insaid host.
 34. The method according to claim 33, wherein said implantingis subcutaneous.
 35. A method for preparing an implantable membranecomprising the steps of: (a) forming a composition comprising adispersion of a second polymer within a matrix of a first polymer, saiddispersion forming a network of microdomains which are notphotomicroscopically observable when hydrated at 400× magnification orless; (b) maintaining said composition at a temperature sufficient tomaintain said first polymer and said second polymer substantiallysoluble; (c) applying said composition at said temperature to asubstrate to form a film thereon; and (d) permitting said resultant filmto dry to form said membrane.
 36. The method of claim 35, wherein saidforming step comprises forming a mixture or a blend.
 37. The method ofclaim 35, wherein said maintaining step comprises heating thecombination to a temperature of about 70° C.
 38. The method of claim 35,wherein said permitting step comprises drying at a temperature fromabout 120° C. to about 150° C.
 39. The method of claim 35, wherein saidsecond polymer is polyethylene oxide.
 40. The method of claim 35,wherein said first polymer is a polyurethane.
 41. The method of claim35, wherein the second polymer is a random or ordered block copolymerselected from the group consisting of ABA block copolymer, BAB blockcopolymer, AB random alternating block copolymer, AB regularlyalternating block copolymer and combinations thereof.
 42. An implantablemembrane formed in accordance with the method of claim 35.